The present invention relates to a fused protein of an adult T cell leukemia virus antigen peptide and an enzyme, a recombinant plasmid wherein a DNA fragment coding for the peptide and the fused protein is incorporated, a microorganism containing the plasmid and a process for producing the adult T cell leukemia virus antigen peptide or the fused protein of the peptide and the enzyme using the microorganism.
Adult T cell leukemia virus (hereinafter referred to as ATLV), which is a synonym of human T cell leukemia virus (HTLV), is a C-type retrovirus isolated from patients with adult T cell leukemia (hereinafter referred to as ATL) [Yoshida, et al., Proc. Natl. Acad. Sci., USA, 79, 2031-2035 (1982)]. There are numerous reports that ATL patients have a poor prognosis and that efficacious treatment does not exist leading to a 50% mortality rate within a half-year.
In recent years, an antibody which reacts specifically with cultured MT-1 cells derived from ATL has been shown to exist in the serum of ATL patients [Hinuma, et al., Proc. Natl. Acad. Sci., USA, 78, 6476-6480 (1980)]. The existence of this antibody has been confirmed subsequently in all ATL patients and the corresponding antigen is called ATL-associated antigen (hereinafter referred to as ATLA). It has been found that the antibody specific for ATLA (hereinafter referred to as Anti-ATLA antibody) exists in 25% of normal, healthy people in areas with a high incidence of ATL. It has also been shown that the distribution of cases possessing the anti-ATLA antibody corresponds to the regions with high ATL incidence (the reference mentioned above). Furthermore, it has been shown that the C-type retrovirus is generated within MT-1 cells, that ATLA is mainly an antigen of this retrovirus and that the anti-ATLA antibody reacts with a structural protein of th virus, particularly the p24 protein. The existence of ATLV genome in MT-1 cells and the peripheral lymphocytes of patients has been established [Yoshida, et al., Proc. Natl. Acad. Sci., USA, 79, 2031-2035 (1982)]. ATLV has also been detected by culturing the lymphocttes of normal people who are positive to the anti-ATLA antibody.
There is a very close correlation between ATL and ATLV, and ATLV is considered to be the causative virus of ATL. Though the route by which infection occurs is still unknown, it is pointed out that coitus transmission and maternal transmission are the most likely routes because of the familial accumulation of the infection.
Further, transfusion of blood is mentioned as an important route. Actually, there has been a report of a clinical example showing that transfusion of blood positive to the anti-ATLA antibody caused the receptor to become positive to the anti-ATLA antibody. As 25% of healthy people in areas with a high incidence of ATL are anti-ATLA antibody positive, the likelihood of their being carriers of ATLV is extremely high, which means that they must be avoided as blood transfusion donors.
The detection of the anti-ATLA antibody is now carried out by the indirect fluorescence antibody method using acetone fixed slides of cultured cells derived from ATL. However, the method is inconvenient for the purpose of analyzing many serum samples rapidly at a time.
In the case of other infectious diseases, it has been known that the detection method using an antigen itself instead of cells is useful. A method using a cell extract of the cultured cells derived from ATL, such as MT-2, as an antigen has been studied. However, the method has the following problems: (1) the culturing of cells is expensive, (2) there is a problem of safety in the mass production of these cells producing ATLV, (3) the amount of the antigen extracted from cells is limited. Therefore, a method for producing ATLA at low cost and in a large amount is still in demand.
The present inventors studied about a method for providing ATLA which is useful for the detection of the anti-ATLA antibody in a large amount and at low cost. As the result, it was found that an ATLV antigen peptide was accumulated in a large amount by culturing a microorganism containing a recombinant DNA which was obtained by incorporating a DNA fragment of gag gene coding for the main antigen peptide p24 in the ATLV genome into a vector DNA by recombinant DNA techniques (Japanese Patent Application No. 170908/83).
The present inventors have further studied for the purpose of more efficient expression. To this end, a recombinant DNA which is useful for the production of a fused protein of an antigen peptide and an enzyme, which is expected to be useful for the simple detection of the antigen peptide and the actual detection of the anti-ATLA antibody, has been prepared and efficient production of said fused protein by a microorganism containing said DNA has now been confirmed.